Malaria has a devastating impact on public health and welfare in the African continent. An important approach for reducing or eliminating malaria impact is vector control. However, vector control is inefficient if not all major vector species are targeted. Mosquitoes of subgenus Cellia - Anopheles gambiae, An. arabiensis, An. funestus, An. moucheti, and An. nili - are the major malaria vectors because they are anthropophilic and susceptible to Plasmodium falciparum. An. gambiae, An. arabiensis, and An. funestus are being intensively studied with the help of cytogenetic and molecular markers. An. moucheti and An. nili have received significantly less attention because the tools needed to precisely characterize species diversity and population structure are lacking. This proposal offers a unique opportunity to develop cytogenetic research tools and to assess their effectiveness for taxonomic and population genetics studies of the neglected malaria vectors An. moucheti and An. nili. The commonly used markers for inferring anopheline taxonomic and population status include chromosomal inversions and microsatellites. Reduced recombination and selection can influence loci within inversions or near inversion breakpoints, resulting in estimates of gene flow that may depart significantly from those based on loci elsewhere in the genome. Therefore, the physical location of microsatellite markers with respect to polymorphic inversions is important information for interpreting population genetic structure. Innovative approaches will be used for developing next-generation-quality cytogenetic and physical maps for malaria vectors. In addition to the traditional cytogenetic and mapping protocols, the project will utilize a novel high-pressure method to prepare polytene chromosome spreads and the primed in situ labeling (PRINS), which is based on a primer-mediated DNA synthesis and used on chromosomes of various organisms. Integrated maps of polytene chromosomes, inversion breakpoints, and microsatellite markers will revolutionize population studies of the neglected malaria vectors. Using these maps, the proposed project will test a hypothesis that there are inversion differences among members of the An. moucheti and An. nili groups. Briefly, the specific aims of this R21 proposal are to 1. Develop high-resolution cytogenetic photomaps for the malaria vectors An. moucheti and An. nili. 2. Determine the taxonomic status and population structure of members of the An. moucheti and An. nili groups using fixed and polymorphic inversions. 3. Identify the chromosomal locations of microsatellite markers for An. moucheti and An. nili with respect to polymorphic inversions. PUBLIC HEALTH RELEVANCE Malaria has a devastating impact on public health and welfare in the African continent. An important approach for reducing or eliminating malaria impact is vector control. This proposal offers a unique opportunity to develop cytogenetic research tools and to assess their effectiveness for taxonomic and population genetics studies of the neglected malaria vectors An. moucheti and An. nili.